It has been recognized that estradiol, i.e. estra-1,3,5(10)-triene-3,17.beta.-diol, and certain analogs and derivatives thereof have estrogen receptor activity. In recent years, it has been postulated that radioiodinated estradiol might be useful for several diagnostic purposes including applications in estrogen receptor assay, radioassay of estrogens and in-vivo radioimaging of certain tumors and organs. As such estradiols and related compounds and particularly radioiodinated analogs thereof have received considerable investigation and publication in the literature. See, for example, Arunachalam et al., J. Biol. Chem., 254, 5900-5 (1979); Tarle et al., J. Lab. Comp. Radiopharm. Vol. XV, pp. 665-671 (1978); Maysinger et al. J. Chromatog. 130, 129-138 to (1977); J. Steroid Biochem. 5, 749-756 (1974), Antar et al., Fed. Proc. 33 (3 Part 1), p. 267(1974); Thaleur et al., Int. J. Appl. Rad. and Isotopes, 27 585-588(1978) Kormai et al., J. Nuclear Med. 18, 360-366 (1977); Albert et al J. Biol. Chem. 177, 247-266 (1949); Ghanadian et al., Int. J. App. Rad. and Isotopes 26, 343-46 (1975); Katzenellenbogen et al., Biochem. 14, 1742-1750 (1975); and Korenman, Steroids 13, 163-177 (1969).
As stated In Katzenellenbogen et al., supra, if an estrogen derivative is to be used in an estrogen receptor assay or for in-vivo radioimaging techniques, it must fulfill three criteria: (1) the linkage of the iodine atom to the ligand must be chemically and metabolically stable; (2) the site of iodine substitution must be such that the derivative is still capable of binding with high affinity to the specific-estrogen binding proteins found in estrogen target tissues, and (3) the physicochemical properties of the derivative (polarity, etc.) must be such that it does not bind excessively to the low affinity, high capacity binding proteins found in serum and tissues. None of the radioiodinated estrogen derivatives studied to date appear to meet the above three requirements necessary for such applications. In addition to the three requirements outlined by Katzenellenbogen et al. above useful radioactive estrogen derivatives should also have high specific activity and be easy to make.
It has been recognized that aliphatic iodine compounds have a high susceptibility toward substitution and elimination reactions and, on the grounds, of chemical stability, it is preferred to have the iodine bonded to an aryl group. Although several compounds having aryl groups substituted with iodine have been made that appear to be metabolically stable, such compounds have shown either low or negligible estrogen receptor activity.
A radioiodinated estrogen analog meeting the three requirements set forth above is highly desired in the medical field.
The chemistry of C-16 halides of estrone methyl ester, including proof of structure, isomerization, interconversion and reduction, has been thoroughly discussed by Mueller et al., J. Am. Chem. Soc. 26, 2403-2413 (1961).
In 1969, Korenman, supra, reported on a study of comparative binding affinity of various estrogens and its relation to estrogenic potency. He concluded that no unitary criterion of relative estrogenic potency can be given. Katzenellenbogen et al., supra, reported that the estrogen receptor is relatively intolerant of bulk at A-ring positions 2 and 4 so that functionalization of these positions would not be consistent with high receptor binding.
Although tritium and carbon 14 remain the usual radioactive isotopes for labeling steroids, for the past several years attempts have been made to use other radioactive isotopes for this purpose. These isotopes include 18-fluorine and 75-selenium, but the commonest have been the radioactive isotopes of iodine. Arunachalam et al., supra, state that so far the results of these endeavors have not been very successful in part due to nonspecific labeling techniques, to loss of biologic activity following iodination, or to nonspecific binding of the iodinated molecule.
Arunachalam et al. undertook to evaluate the utility of iodoestradiol analogs made highly radioactive with iodine isotopes in (a) the non-invasive differentiation of estrogen-dependent from estrogen-independent breast tumors, (b) spread of metastases containing estrogen receptors, and (C) potential application in therapeutic irradiation of target tissues. The model syntheses of a number of nonradioactive .sup.127 I-estrogen analogs are described. The analogs were tested for their ability to displace (compete with) [.sup.3 H]estradiol from receptor sites. The most active compounds, 16.beta.-iodoestra-1,3,5(10)-triene-3-17.beta.-diol and 6-iodoestra-1,3,5(10), 6-tetraene-3,17.beta.-diol showed a relative binding affinity of 0.57 and 0.49, respectively. They prepared and evaluated 16.alpha.-iodo-3-hydroxyestra-1,3,5(10)-triene-17-one but concluded the introduction of the iodine group at the 16.beta. position does not diminish binding to the same extent as 16.alpha. substitution and stated that this finding was consistant with a report on the difference in binding between estriol and epiestriol. Further, Counsell et al., in Steroid Hormone Action and Cancer, Menon and Reel, Eds., Plenum, New York (1976) at p. 107 indicate that the 16.alpha. substituted estrone would not be useful. This is probably because the estrone is metabolically and chemically unstable.
Upon considering these disclosures, one skilled in the art would not contemplate that iodo substitution at the 2,4, or 16.alpha. position would fulfill the three requirements set forth above.